The proposed K99/R00 application incorporates a comprehensive research and training plan for studying ion channels and channelopathies in the olfactory system. Ion channels are critical for regulating excitability in many cell types includin olfactory sensory neurons (OSNs). These proteins regulate the movement of ions across the cellular membrane. In OSNs ion channels are responsible for depolarizing the cell in response to odor stimulation, initializing an action potential and synaptic transmission. Channelopathies are a class of human genetic disorders in which ion channel function is disrupted leading to defects in multiple organ systems. Disruptions in ion channels are known to cause epilepsy, cardiac arrhythmias, blindness, deafness and alterations in pain sensitivity. Channelopathies in human patients can result from both loss-of- function and gain-of-function mutations in ion channel genes. It has recently been shown that channelopathies can result in anosmia in humans. Deletions of several different ion channels in mouse models also causes anosmia, indicating that olfactory signaling can be affected at multiple steps. The proposed research will analyze the ability of gene therapy to correct channelopathy induced defects in olfactory function at these different stages in signaling. Aim 1 of this proposal will use two mouse strains with targeted deletions in cyclic nucleotide gated (CNG) channel subunits. CNGA2 and CNGB1 are critical subunits of the olfactory CNG channel necessary for odor detection and their loss leads to anosmia. Using adenovirus, I will deliver functional copies of the missing gene to mutant OSNs to test the ability to restore olfactory function to anosmic animals. Restoration of the sense of smell will be analyzed with electrophysiological and behavioral methods. Aim 2 will investigate the ability of gene therapy to correct channelopathy induced defects in synaptic transmission due to loss of the sodium channel alpha-subunit Nav1.7 (encoded by Scn9a) in OSNs. Using olfactory specific Scn9a null mice, I will use adenovirus to express ectopic Scn9a in OSNs and analyze the restoration of synaptic transmission. In addition this aim will analyze the effects of gain-of-function mutations in Scn9a through adenovirus-mediated expression of identified mutant sodium channels in OSNs. This will test the effect of hyper-excitability on olfactory function. Finally Aim 3 will analyze the role of two voltage gated calcium channel (VGCC) alpha-subunits, Cav1.3 and Cav2.2, in olfactory function. Mutations in VGCCs, including CACNA1D (encoding Cav1.3), underlie channelopathy disorders affecting sensory function and are therefore potential causes of anosmia. An understanding of the function of these VGCCs in the olfactory system will help to direct the identification of novel mutations in anosmic patients. Together the results from these 3 aims will provide new insight into the mechanisms of olfactory signaling and the ability of gene therapy to correct defects in ion channel function. The results from the proposed research will be important for helping to develop therapies for patients with anosmia due to channelopathies. In addition these results may provide insight into developing treatments for other sensory defects such as vision and hearing loss.